CN106674197A - Aurora kinase A inhibitor and preparation and application thereof - Google Patents
Aurora kinase A inhibitor and preparation and application thereof Download PDFInfo
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- CN106674197A CN106674197A CN201510753988.3A CN201510753988A CN106674197A CN 106674197 A CN106674197 A CN 106674197A CN 201510753988 A CN201510753988 A CN 201510753988A CN 106674197 A CN106674197 A CN 106674197A
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- C07—ORGANIC CHEMISTRY
- C07D—HETEROCYCLIC COMPOUNDS
- C07D401/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom
- C07D401/02—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings
- C07D401/12—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings linked by a chain containing hetero atoms as chain links
Abstract
The invention discloses a novel stable nitroxide radical-marked aurora kinase A inhibitor and a preparation method and application thereof. The aurora kinase A inhibitor is N-(5-fluoro-4-o-chloroaniline-pyrimidin-2)-aminobenzoic acid 4-amino-2, 2, 6, 6-tetramethylpiperidyl oxynitride amide, or N-{5-fluoro-4-amino-(benzoyl o-chloroaniline)-pyrimidin-2}-aminobenzoic acid 4-amino-2, 2, 6, 6-tetramethylpiperidyl oxynitride amide. According to the preparation method, a target compound is prepared by performing a substitution reaction on 2, 4-dichloro-5-substituted-pyrimidine and aniline or p-aminobenzanilide and then 4-amino-2, 2, 6, 6-tetramethylpiperidyl nitrogen oxide. The aurora kinase A inhibitor is applied to preparation of anticancer medicines.
Description
Technical field
The present invention relates to organic chemistry and medicinal chemistry art, specially a kind of amidos of 2,4- bis- of stable nitrogen-oxygen free radical mark are phonetic
Pyridine class BTAK inhibitor, and the preparation method and purposes of this inhibitor.
Background technology
Laser kinases (Aurora kinase) is the new threonine/serineprotein kinase of a class, at centerbody duplication, the two poles of the earth
Vital work is played during the important mitosis such as spindle formation, chromosomal rearrangement and the monitoring of chromosome examination point
With (Cancer Metastasis Rev.2003,22,451).There is the Asia of three kinds of 26S Proteasome Structure and Function height correlations in Aurora A family
Group:That is Aurora-A, Aurora-B and Aurora-C.Their prlmary structure of protein contains a N- ends control region and
Individual C- ends catalytic domain, and enzyme domains sequence similarity is also identical up to the residue of 71%, ATP adenine ring binding sites；But
They have entirely different and non-overlapping copies functions in cell division.Aurora A are from the mitotic S end of term to next point
Split the G phases in cycle, affect the separation of centerbody, the formation of ripe and the two poles of the earth spindle (Nat.Rev.Cancer 2005,
5,42).Aurora B are located at the centromere region of chromosome in mitosis early stage, and anaphase then moves to micro- from centromere
Pipe.It is relatively fewer to the research of Aurora C functions at present.1998, American scholar Bischoff etc. found first Aurora
Kinases is over-expressed in various cancer cells, and the process such as the unstability with chromosome, canceration, tumor proliferation and chemoresistance
Closely related (EMBO is J.1998,17,3052).Due to the unique pharmacological mechanism of Aurora A, with Aurora A
For target spot drug development become cancer therapy drug research one of focus (Expert Opin.Drug Discovery 2011,6,
291)。
So far, also it is used for clinical treatment without Aurora A inhibitor, only several Aurora A inhibitor are being carried out
Clinical testing, such as VX-680, PHA-739358, AZD-1152, MLN8054, SNS-314, ENMD-2076 (medicine
Learn progress 2008,32,337；Chinese antibiotic magazine 2010,35 (9), 641).But, these small molecules are all existed not
With the toxic and side effect of degree, although drug therapy has a bright future, it need further chemical modification research, reduces its side effect.
For Aurora-B, Aurora-A is excessive in many tumours such as colon cancer, oophoroma, cancer of the stomach and breast cancer
Expression.The overexpression of Aurora-A makes the monitoring function of cell mitogen test point out of control, and cause a split exception, oncogene
Amplification, normal cell turnover be cancer cell (J.Cell Sci.2007,120,2987).Aurora-A can more efficiently cause cell
Mitotic arrest and rapid induction Apoptosis, thus, the antineoplastic with Aurora-A as target spot is treated to a certain degree
On will be better than Aurora-B (Curr.Opin.Invest.Drugs 2006,7,1044).The selective Aurora-A kinases just reported
For inhibitor, the MLN8237 that at present only Millennium companies of Canada develop is carrying out I clinical trial phase (Proc.Natl.
Acad.Sci.USA 2007,104,4106).Recently, American scholar Cochran and Lwerence report respectively a class 2,4- bis-
The cyclosubstituted single pyrimidine cyclics of benzene, some compounds are up to 1000 times to the selectivity of Aurora-A compared with Aurora-B, but
Because their dissolubility and/or film penetration capacity are poor, cause its antitumor activity relatively low, lose the value of further exploitation
(J.Med.Chem.2012,55,7392).In addition, the phthalein that a class novel pyrazole of Norway's scholar Prime M.E. reports replaces
Zionoes compound, the selectivity to Aurora-A kinase inhibitions is more than 1000, whether enters preclinical study at present unclear
(J.Med.Chem.2011,54,312)。
In view of stable nitrogen-oxygen free radical as effective drug transporters, the dissolubility of medicine can be improved, and promote medicine preferential
Through cancerous tumor cell film (Science 2001,291,266；J.Med.Chem.2005,48,6393).Present invention design synthesis one
The pyrimidines of stable nitrogen-oxygen free radical mark are planted, experiment in vitro shows that this compound has preferable cell to cancer cell
Toxicity, and to Aurora A compared with the selective inhibitory action of Aurora B, while this compound has appropriate fat moisture disposition
Number is a kind of with the compound that exploitation is new anticancer drug；In preparation method, the present invention passes through straightforward procedure efficiently
Key intermediate and its target compound are synthesized.
The content of the invention
It is an object of the invention to by the use of stable nitrogen-oxygen free radical as effective drug transporters, expect the dissolving for improving compound
Property, and promote compound preferentially through cancerous tumor cell film, play anticancer effect.Cytotoxicity experiment finds the compounds of this invention pair
Various cancer cells have stronger In-vitro Inhibitory Effect, while to Aurora A compared with the selective inhibitory action of Aurora B.
A kind of BTAK inhibitor of the stable nitrogen-oxygen free radical mark of the structure with active anticancer such as formula I,
Wherein：
R is ortho position chlorine, or meta chlorine, or contraposition chlorine, or vicinal hydroxyl groups, or meta-hydroxyl, or contraposition hydroxyl.
X is nitro, or fluorine, or chlorine, or bromine.
A kind of BTAK inhibitor of the stable nitrogen-oxygen free radical mark of the structure with active anticancer such as formula II,
Wherein：
R is ortho position chlorine, or meta chlorine, or contraposition chlorine.
X is nitro, or fluorine, or chlorine, or bromine.
The preparation method of compound shown in Chinese style of the present invention I, is characterized in that with the chloro- 5- nitro-pyrimidines of 2,4- bis-, or the chloro- 5- of 2,4- bis-
Fluoropyrimidine, or 2,4,5- trichloropyrimidines, or the chloro- 5- Bromopyrimidines of 2,4- bis- are raw material, first and o-chloraniline, or m-chloroaniline, or
Parachloroanilinum, or ortho-aminophenol, or a hydroxyanilines, or the reaction of para hydroxybenzene amine to generate the chloro- 4- o-chloranilines -5- nitros of 2- phonetic
Pyridine, or the chloro- 4- m-chloroanilines -5- nitro-pyrimidines of 2-, or the chloro- 4- parachloroanilinum -5- nitro-pyrimidines of 2-, or the chloro- 4- ortho-aminophenols of 2-
Hydroxyanilines -5- nitro-pyrimidines between -5- nitro-pyrimidines, or 2- chloro- 4-, or the chloro- 4- para hydroxybenzenes amine -5- nitro-pyrimidines of 2-, or 2- chlorine
- 4- o-chloranilines -5-FU, or the chloro- 4- o-chloranilines pyrimidine of 2,5- bis-, or the chloro- 4- o-chloranilines -5- Bromopyrimidine (J.Med. of 2-
Chem.2012,55,7392)；While with the p-aminobenzoic acid of tertbutyloxycarbonyl protection as raw material, in 1- (3- dimethylaminos third
Base) -3- ethyl-carbodiimide hydrochlorides (EDCI) and I-hydroxybenzotriazole (HOBT) exist under conditions of with 4- amino -2,2,6,6-
Tetramethyl piperidine nitrogen oxides (TMPO) is condensed, then deprotection base is obtained final product to ammonia in the dichloromethane solution of trifluoroacetic acid
Base benzoyl 4- amino -2,2,6,6- tetramethyl piperidine nitrogen oxides (J.Med.Chem.2001,44,441)；It is last obtained above
Two kinds of compounds obtain the target compound representated by formula I under strong acid catalyst in the 2- positions generation substitution reaction of pyrimidine ring.
The preparation method of compound shown in Chinese style of the present invention II, is characterized in that with the chloro- 5- nitro-pyrimidines of 2,4- bis-, or the chloro- 5- of 2,4- bis-
Fluoropyrimidine is raw material, first with p-benzoyl o-chloraniline, or p-benzoyl m-chloroaniline, or p-benzoyl
Parachloroanilinum reaction generation N- (the chloro- 5- nitro-pyrimidins -4 of 2-)-aminobenzoyl o-chloraniline, or N- (the chloro- 5- nitro-pyrimidins -4 of 2-) -
Aminobenzoyl m-chloroaniline, or N- (the chloro- 5- nitro-pyrimidins -4 of 2-)-aminobenzoyl parachloroanilinum, or N- (the fluoro- pyrimidines of the chloro- 5- of 2-
- 4)-aminobenzoyl o-chloraniline (J.Med.Chem.2012,55,7392)；The p-aminophenyl protected with tertbutyloxycarbonyl simultaneously
Formic acid is raw material, in 1- (3- dimethylamino-propyls) -3- ethyl-carbodiimide hydrochlorides (EDCI) and I-hydroxybenzotriazole
(HOBT) it is condensed with 4- amino-2,2,6,6-tetramethylpiperidine nitrogen oxides (TMPO) under conditions of existing, then in trifluoro second
Deprotection base obtains final product p-benzoyl 4- amino -2,2,6,6- tetramethyl piperidine nitrogen oxides (J. in the dichloromethane solution of acid
Med.Chem.2001,44,441)；Last two kinds of compounds obtained above take under strong acid catalyst in the 2- positions of pyrimidine ring
Generation reaction obtains the target compound representated by formula II.
The preparation method of the compound described in Chinese style of the present invention I, II, is characterized in that p-benzoyl 4- amino in reaction
The amount of substance of -2,2,6,6- tetramethyl piperidines nitrogen oxides and corresponding N- (the chloro- 5- replacement-pyrimidines -4 of 2-)-amino benzoyl chloride aniline reaction
Than for 1:1.05-1.20, reaction temperature is 70-110 DEG C, and catalyst is hydrochloric acid.
Compound described in Chinese style of the present invention I, II is to human cervical carcinoma cell HeLa, Non-small cell lung carcinoma cell A-549, people
Tetra- kinds of growth of cancer cells of hepatocellular carcinoma H22 and human colon cancer cell LoVo have significant inhibitory action, wherein the overwhelming majorityization
Compound is significantly stronger than positive control VX-680 to the inhibitory action of these cancer cell multiplications, especially chemical compounds I g and II d to this four
Planting cancer cell has stronger activity, and BTAK (Aurora A) is shown compared with aurora kinase B (Aurora B)
Obvious Selective depression.Therefore the compound of the present invention can be used to prepare cancer therapy drug.
The method of the present invention is further illustrated below by embodiment.It should be understood that the preparation method of the embodiment of the present invention is only
It is the letter for illustrating the present invention, rather than limitation of the present invention, under the concept thereof of the present invention to preparation method of the present invention
Single improvement belongs to the scope of protection of present invention.
Specific embodiment
The present invention provides below example：
Embodiment 1：The preparation of the chloro- 4- o-chloranilines -5- nitro-pyrimidines of 2-
Take the chloro- 5- nitro-pyrimidines (205mg, 1.05mmol) of 2,4- bis- and be dissolved in dry dichloromethane, adjacent chlorobenzene is added under room temperature
Amine (125mg, 1.0mmol) simultaneously continues to react 8h, and reaction is evaporated off solvent after terminating, and the direct column chromatography of residue obtains product 2-
Chloro- 4- o-chloranilines -5- nitro-pyrimidines.Yield:89%；Yellow solid；1H-NMR(600MHz,CDCl3)δ10.68(s,1H),
9.22(s,1H),8.34-8.32(m,1H),7.51-7.21(m,3H)。
With m-chloroaniline, or parachloroanilinum, or ortho-aminophenol, or a hydroxyanilines, or para hydroxybenzene amine replaces respectively adjacent chlorine
Aniline reaction prepares the chloro- 4- m-chloroanilines -5- nitro-pyrimidines of 2-, or the chloro- 4- parachloroanilinum -5- nitro-pyrimidines of 2-, or the chloro- 4- neighbours hydroxyls of 2-
Base aniline -5- nitro-pyrimidines, or hydroxyanilines -5- nitro-pyrimidines between 2- chloro- 4-, or the chloro- 4- para hydroxybenzenes amine -5- nitro-pyrimidines of 2-.
With the chloro- 5-FUs of 2,4- bis-, or 2,4,5- trichloropyrimidines, or the chloro- 5- Bromopyrimidines of 2,4- bis- replace the chloro- 5- nitro-pyrimidines of 2,4- bis-
Reaction prepares the chloro- 4- o-chloranilines -5-FUs of 2-, or the chloro- 4- o-chloranilines pyrimidine of 2,5- bis-, or the chloro- 4- o-chloranilines -5- bromines of 2-
Pyrimidine.
Embodiment 2：The preparation of p-aminobenzoic acid 4- amino -2,2,6,6- tetramethyl piperidine nitrogen oxides acid amides
The p-aminobenzoic acid for taking 1.8g Boc protections is dissolved in dry dichloromethane, sequentially adds 1.8g EDC, 1.22g
HOBt and 1.53g compound TMPO.It is stirred at room temperature complete to reaction.Organic phase adds water washing, and dry with anhydrous magnesium sulfate
It is dry；Solvent and column chromatography are evaporated off, pink solid 2.52g, yield 90% is obtained.MS(ESI)392.15[M+2H]+。
Weigh above-mentioned reaction products therefrom 2.5g to be dissolved in 15ml dichloromethane, 6ml trifluoroacetic acids are added dropwise, reaction is stirred at room temperature
After 0.5h, solvent is removed under reduced pressure, gained solid column chromatography obtains white solid 1.74g, yield 94%.MS(ESI)292.15
[M+2H]+。
Embodiment 3：N- (5- nitros -4- o-chloranilines-pyrimidine -2)-aminobenzoic acid 4- amino -2,2,6,6- tetramethyl piperidine nitrogen oxides acid amides
Preparation
By the chloro- 4- o-chloranilines -5- nitro-pyrimidines (280mg, 1mmol) of 2- and p-benzoyl 4- amino -2,2,6,6- tetramethyls
During piperidine nitroxide (430mg, 1.5mmol) is dissolved in ethanol, dioxane solution 2ml of hydrochloric acid is added, be heated to reflux.
Question response substantially completely stops afterwards reaction, and solvent is evaporated off, and direct column chromatography obtains yellow solid, yield:62%.
m.p.:208-210℃；HPLC:MeOH:H2O (0.1%TFA)=65:35,0.8ml/min, t=21.23min, 95.0%；IR
(KBr,cm-1)3362,3015,1629,1536,1506,1422,1332,1195,1025,836,726；ESR(DMSO):G=
2.006, An (G)=15.96, △ H (G)=2.95；HRMS(ESI)540.2111for[M+2H]+(calcd 540.2121for
C26H31N7O4Cl).
In carrying out aftermentioned cell proliferation Inhibition test, the present embodiment sample number into spectrum is I a.
Embodiment 4：N- (5- nitros -4- m-chloroanilines-pyrimidine -2)-aminobenzoic acid 4- amino -2,2,6,6- tetramethyl piperidine nitrogen oxides acid amides
Preparation
Operating process is same with case study on implementation 3, simply replaces the chloro- 4- o-chloranilines -5- of 2- with the chloro- 4- m-chloroanilines -5- nitro-pyrimidines of 2-
Nitro-pyrimidine, obtains yellow solid, yield:68%.
m.p.:217-219℃；HPLC:MeOH:H2O (0.1%TFA)=72:28,0.8mL/min, t=13.70min, 98.0%；IR
(KBr,cm-1)3376,3281,3022,1636,1610,1584,1541,1526,1481,1414,1329,1202,1137,858,
726；ESR(DMSO):G=2.006, An (G)=15.92, △ H (G)=2.75；HRMS(ESI)540.2117for
[M+2H]+(calcd 540.2121for C26H31N7O4Cl).
In carrying out aftermentioned cell proliferation Inhibition test, the present embodiment sample number into spectrum is I b.
Embodiment 5：N- (5- nitros -4- parachloroanilinum-pyrimidine -2)-aminobenzoic acid 4- amino -2,2,6,6- tetramethyl piperidine nitrogen oxides acid amides
Preparation
Operating process is same with case study on implementation 3, simply replaces the chloro- 4- o-chloranilines -5- of 2- with the chloro- 4- parachloroanilinum -5- nitro-pyrimidines of 2-
Nitro-pyrimidine, obtains yellow solid, yield:72%.
m.p.:225-227℃；HPLC:MeOH:H2O (0.1%TFA)=72:28,0.8mL/min, t=14.18min, 96.5%；IR
(KBr,cm-1)3379,2995,1639,1616,1579,1529,1489,1404,1202,841,719；ESR(DMSO):G=
2.006, An (G)=15.88, △ H (G)=2.85；HRMS(ESI)540.2117for[M+2H]+(calcd 540.2121for
C26H31N7O4Cl).
In carrying out aftermentioned cell proliferation Inhibition test, the present embodiment sample number into spectrum is I c.
Embodiment 6：N- (5- nitros -4- ortho-aminophenols-pyrimidine -2)-aminobenzoic acid 4- amino -2,2,6,6- tetramethyl piperidine nitrogen oxides acyls
The preparation of amine
Operating process is same with case study on implementation 3, simply replaces the chloro- 4- o-chloranilines -5- of 2- with the chloro- 4- ortho-aminophenols -5- nitro-pyrimidines of 2-
Nitro-pyrimidine, obtains yellow solid, yield:61%.
m.p.:126-128℃；HPLC:MeOH:H2O (0.1%TFA)=80:20,0.8mL/min, t=7.18min, 95.6%；IR
(KBr,cm-1)3369,2940,1641,1624,1576,1411,1459,1327,1205,838,746；ESR(DMSO):G=
2.006, An (G)=15.96, △ H (G)=2.95；HRMS(ESI)522.2463for[M+2H]+(calcd 522.2459for
C26H32N7O5).
In carrying out aftermentioned cell proliferation Inhibition test, the present embodiment sample number into spectrum is I d.
Embodiment 7：N- (hydroxyanilines-pyrimidine -2 between 5- nitro -4-)-aminobenzoic acid 4- amino -2,2,6,6- tetramethyl piperidine nitrogen oxides acyls
The preparation of amine
Operating process is same with case study on implementation 3, simply replaces the chloro- 4- o-chloranilines -5- of 2- with hydroxyanilines -5- nitro-pyrimidines between 2- chloro- 4-
Nitro-pyrimidine, obtains yellow solid, yield:52%.
m.p.:97-99℃；HPLC:MeOH:H2O (0.1%TFA)=77:23,0.8mL/min, t=8.41min, 99.8%；IR(KBr,
cm-1)3324,2985,1678,1619,1591,1529,1422,1332,1207,853,786,724；ESR(DMSO):G=
2.006, An (G)=15.86, △ H (G)=2.98；HRMS(ESI)522.2453for[M+2H]+(calcd 522.2459for
C26H32N7O5).
In carrying out aftermentioned cell proliferation Inhibition test, the present embodiment sample number into spectrum is I e.
Embodiment 8：N- (5- nitro -4- para hydroxybenzene amine-pyrimidine -2)-aminobenzoic acid 4- amino -2,2,6,6- tetramethyl piperidine nitrogen oxides acyls
The preparation of amine
Operating process is same with case study on implementation 3, simply replaces the chloro- 4- o-chloranilines -5- of 2- with the chloro- 4- para hydroxybenzenes amine -5- nitro-pyrimidines of 2-
Nitro-pyrimidine, obtains yellow solid, yield:63%.
m.p.:234-236℃；HPLC:MeOH:H2O (0.1%TFA)=68:32,0.6mL/min, t=8.68min, 98.5%；IR
(KBr,cm-1)3421,3281,2344,1686,1616,1579,1499,1394,1267,1200,841,744,699；ESR
(DMSO):G=2.006, An (G)=15.94, △ H (G)=3.05；HRMS(ESI)522.2454for[M+2H]+(calcd
522.2459for C26H32N7O5).
In carrying out aftermentioned cell proliferation Inhibition test, the present embodiment sample number into spectrum is I f.
Embodiment 9：N- (the fluoro- 4- o-chloranilines-pyrimidines -2 of 5-)-aminobenzoic acid 4- amino -2,2,6,6- tetramethyl piperidine nitrogen oxides acid amides
Prepare
Operating process is same with case study on implementation 3, simply replaces the chloro- 4- o-chloranilines -5- nitre of 2- with the chloro- 4- o-chloranilines -5-FUs of 2-
Yl pyrimidines, obtain white solid, yield:50%.
m.p.:137-139℃；HPLC:MeOH:H2O (0.1%TFA)=50:50,0.8mL/min, t=34.94min, 95.8%；IR
(KBr,cm-1)3384,3277,2980,2942,2628,2496,1678,1606,1574,1511,1494,1444,1399,1327,
1190,1038,850,776；ESR(DMSO):G=2.006, An (G)=15.92, △ H (G)=2.85；HRMS(ESI)
512.2090for[M+H]+(calcd 512.2097for C26H31N6O2FCl).
In carrying out aftermentioned cell proliferation Inhibition test, the present embodiment sample number into spectrum is I g.
Embodiment 10：N- (the chloro- 4- o-chloranilines-pyrimidines -2 of 5-)-aminobenzoic acid 4- amino -2,2,6,6- tetramethyl piperidine nitrogen oxides acid amides
Preparation
Operating process is same with case study on implementation 3, simply replaces the chloro- 4- o-chloranilines -5- nitre of 2- with the chloro- 4- o-chloranilines -5- chlorine pyrimidines of 2-
Yl pyrimidines, obtain white solid, yield:42%.
m.p.:156-158℃；HPLC:MeOH:H2O (0.1%TFA)=50:50,0.8mL/min, t=32.93min, 100%；IR
(KBr,cm-1)3381,3273,2980,2606,2501,1678,1611,1569,1501,1447,1329,1187,1033,853,
776,751；ESR(DMSO):G=2.006, An (G)=15.90, △ H (G)=2.65；HRMS(ESI)513.1928for
[M-14]+(calcd 527.1729for C26H29N6O2Cl).
In carrying out aftermentioned cell proliferation Inhibition test, the present embodiment sample number into spectrum is I h.
Embodiment 11：N- (the bromo- 4- o-chloranilines-pyrimidines -2 of 5-)-aminobenzoic acid 4- amino -2,2,6,6- tetramethyl piperidine nitrogen oxides acid amides
Preparation
Operating process is same with case study on implementation 3, simply replaces the chloro- 4- o-chloranilines -5- nitre of 2- with the chloro- 4- o-chloranilines -5- Bromopyrimidines of 2-
Yl pyrimidines, obtain white solid, yield:46%.
m.p.:167-168℃；HPLC:MeOH:H2O (0.1%TFA)=65:35,0.8mL/min, t=11.36min, 99.3%；IR
(KBr,cm-1)3366,2982,2940,2499,1609,1569,1509,1494,1444,1387,1324,1187,1033,851,
768；ESR(DMSO):G=2.006, An (G)=15.96, △ H (G)=2.95；HRMS(ESI)557.1422for[M-14]+
(calcd 571.1224for C26H29N6O2ClBr).
In carrying out aftermentioned cell proliferation Inhibition test, the present embodiment sample number into spectrum is I i.
Embodiment 12：N- { 5- nitro -4- amino-(benzoyl o-chloraniline)-pyrimidine -2 }-aminobenzoic acid 4- amino -2,2,6,6- tetramethyl piperazines
The preparation of pyridine nitrogen oxides acid amides
Operating process is same with case study on implementation 3, simply replaces 2- chlorine with the chloro- 5- nitros -4- amino of 2--(benzoyl o-chloraniline)-pyrimidine
- 4- o-chloraniline -5- nitro-pyrimidines, obtain yellow solid, yield:72%.
m.p.:274-276℃；HPLC:MeOH:H2O (0.1%TFA)=75:25,0.8mL/min, t=9.47min, 99.7%；IR
(KBr,cm-1)3404,2995,1656,1631,1539,1424,1332,1222,853,766；ESR(DMSO):G=2.006,
An (G)=15.96, △ H (G)=2.92；HRMS(ESI)659.2490for[M+2H]+(calcd 659.2492for
C33H36N8O5Cl).
In carrying out aftermentioned cell proliferation Inhibition test, the present embodiment sample number into spectrum is II a.
Embodiment 13：N- { 5- nitro -4- amino-(benzoyl m-chloroaniline)-pyrimidine -2 }-aminobenzoic acid 4- amino -2,2,6,6- tetramethyl piperazines
The preparation of pyridine nitrogen oxides acid amides
Operating process is same with case study on implementation 3, simply replaces 2- chlorine with the chloro- 5- nitros -4- amino of 2--(benzoyl m-chloroaniline)-pyrimidine
- 4- o-chloraniline -5- nitro-pyrimidines, obtain yellow solid, yield:59%.
m.p.:248-249℃；HPLC:MeOH:H2O (0.1%TFA)=75:25,0.8mL/min, t=7.96min, 98.7%；IR
(KBr,cm-1)3394,2980,2364,1668,1624,1531,1427,1334,1220,851,761；ESR(DMSO):G=
2.006, An (G)=15.92, △ H (G)=2.90；HRMS(ESI)659.2507for[M+2H]+(calcd 659.2492for
C33H36N8O5Cl).
In carrying out aftermentioned cell proliferation Inhibition test, the present embodiment sample number into spectrum is II b.
Embodiment 14：N- { 5- nitro -4- amino-(benzoyl parachloroanilinum)-pyrimidine -2 }-aminobenzoic acid 4- amino -2,2,6,6- tetramethyl piperazines
The preparation of pyridine nitrogen oxides acid amides
Operating process is same with case study on implementation 3, simply replaces 2- chlorine with the chloro- 5- nitros -4- amino of 2--(benzoyl parachloroanilinum)-pyrimidine
- 4- o-chloraniline -5- nitro-pyrimidines, obtain yellow solid, yield:65%.
m.p.:275-277℃；HPLC:MeOH:H2O (0.1%TFA)=75:25,0.8mL/min, t=9.80min, 97.7%；IR
(KBr,cm-1)3401,2980,2379,1609,1536,1504,1414,1329,1210,853,788；ESR(DMSO):G=
2.006, An (G)=15.94, △ H (G)=2.92；HRMS(ESI)659.2495for[M+2H]+(calcd 659.2492for
C33H36N8O5Cl).
In carrying out aftermentioned cell proliferation Inhibition test, the present embodiment sample number into spectrum is II c.
Embodiment 15：N- { the fluoro- 4- amino of 5--(benzoyl o-chloraniline)-pyrimidine -2 }-aminobenzoic acid 4- amino -2,2,6,6- tetramethyl piperidines
The preparation of nitrogen oxides acid amides
Operating process is same with case study on implementation 3, simply replaces the chloro- 4- of 2- with the fluoro- 4- amino of the chloro- 5- of 2--(benzoyl o-chloraniline)-pyrimidine
O-chloraniline -5- nitro-pyrimidines, obtain white solid, yield:51%.
m.p.:213-215℃；HPLC:MeOH:H2O (0.1%TFA)=60:40,0.8mL/min, t=21.46min, 95.6%；IR
(KBr,cm-1)3424,2962,2925,2850,1614,1594,1509,1424,1322,1232,1182,848,759；ESR
(DMSO):G=2.006, An (G)=15.96, △ H (G)=2.97；HRMS(ESI)632.2542for[M+2H]+(calcd
632.2547for C33H36N7O3FCl).
In carrying out aftermentioned cell proliferation Inhibition test, the present embodiment sample number into spectrum is II d.
Embodiment 16：In vitro cytotoxic effect of the amine pyrimidine class compounds of 2,4- bis- of stable nitrogen-oxygen free radical mark to cancer cell
Synthesized target compound suppresses the ability of tumor cell proliferation in order to study this experiment, and we determine compound
To four kinds of human tumor cells (human cervical carcinoma cell HeLa, Non-small cell lung carcinoma cell A-549, human liver cancer cell HepG2
With human colon cancer cell LoVo) in vitro cytotoxic effect, and using VX-680 as positive control.The detection side that experiment is adopted
Method is the mtt assay of standard.Experimental technique is：
Take the logarithm the cell in growth period, cell suspension is made with the culture mediums of RPMI 1640 containing 10% hyclone, per hole 6000
Individual cell is inoculated into 96 orifice plates, and flat board is put into 37 DEG C, containing 5%CO2The incubator culture 24h of air and 100% humidity makes
Its is adherent, and rear substitution contains the culture mediums of RPMI 1640 (200 μ L/ holes) containing 10% hyclone of variable concentrations medicine, medicine
Concentration is respectively 10-4, 2 × 10-5, 4 × 10-6, 8 × 10-7, 1.6 × 10-7,3.2×10-8Mol/L, and zeroing hole is set, blank group, sun
Property control group VP-16, every group of three wells, incubation culture 48 hours after take out, per hole add 20 μ L MTT (5mg/mL),
Culture 4h being incubated again, making MTT be reduced to Jia Za, suction out supernatant, 150 μ L DMSO are added per hole, concussion Shi Jia Za is brilliant
Body dissolves, and with ELIASA OD value of the cell liquid at 570nm is measured, and calculates the inhibiting rate of each concentration of compound.By inhibiting rate
Calculate IC50Value, and take the mean value of three tests.
Chemical compounds I a-i, II a-d are to human cervical carcinoma cell HeLa, Non-small cell lung carcinoma cell A-549, human liver cancer cell
The in-vitro pharmacological experiments that tetra- kinds of cancer cell multiplications of HepG2 and human colon cancer cell LoVo suppress the results are shown in Table 1.
Chemical compounds I a-the i of table 1, the cytotoxicity (IC of II a-b and VX-68050,μM)
Note：(1) experimental result is the statistics of three parallel laboratory tests；(2) action time：48 hours
Experiment in vitro proves, 13 compounds of synthesis are to HeLa, A-549, HepG2 and LoVo all than positive control VX-680
There is preferably external inhibitory activity；Especially compound ii d shows highest inhibitory activity to these four cancer cells.
Embodiment 16：The selective inhibitory activity of chemical compounds I g and II d to laser kinases A
Laser kinases (Aurora kinase) is in centerbody duplication, the formation of the two poles of the earth spindle, chromosomal rearrangement and chromosome examination point
Vital effect is played during the important mitosis such as monitoring.Aurora A is over-expressed in various cancer cells,
And it is closely related with the process such as the unstability of chromosome, canceration, tumor proliferation and chemoresistance, it is current cancer therapy drug research
One of important target.In order to study the mechanism of chemical compounds I g and the antitumor proliferation functions of II d, while whether may be used to explore it
To suppress the expression of Aurora A in tumour cell as other miazines inhibitor, we adopt enzyme-linked immunosorbent assay
(ELISA) test is studied the inhibitory activity of Aurora A in HepG2 cells.
Concrete grammar：Take the HepG2 cells in exponential phase to be placed in microtiter plate, add in each hole different dense
The medicine of degree gradient, per the μ L samples of hole 10, the sample diluting liquid of 40 μ L, blank well only adds sample, dilution to be placed in 37 DEG C
Insulating box, is incubated 30min, is then washed with cleaning solution 5 times, after patting dry, plus enzyme labelled antibody, per the μ L (blank wells of hole 50
It is not added with) 37 DEG C of insulating box incubation 30min are placed in again.Most backward each μ L tmb substrate liquid of Kong Zhongjia 50,37 DEG C of insulating boxs are incubated
15～30min is educated, is taken out in backward every hole and is added terminate liquid to carry out terminating reaction.Afterwards cell dissociation is centrifuged, then PBS bufferings
Liquid is resuspended, multigelation 3 times, and 20～30min is centrifuged in 2000～3000r/min, and Aspirate supernatant is detected at 450nm
OD values.And thus calculate compound inhibiting rate and IC50Value, and take the mean value of three tests.
Experimental result is shown in Table 2.
Inhibitory activity of the chemical compounds I g of table 2 and II d to laser kinases A and B
Note：(1) experimental result is the statistics of three parallel laboratory tests；
From table 2, chemical compounds I g and II d is more higher than VX 680 to the inhibitory activity of Aurora A, respectively VX 680
20 times and 32 times.Meanwhile, chemical compounds I g and II d have more significantly inhibitory activity to Aurora A than Aurora B,
Wherein effects of the chemical compounds I g to Aurora A is 11 times to Aurora B, and effects of II d to Aurora A is to Aurora
67 times of B, and VX 680 is no to Aurora A and Aurora B selective.
Such compound synthesis method is simple, and raw material is cheap and easy to get, and pharmacologically active significantly, is expected to become and possesses China's independent intellectual
The newtype drug of one class treating cancer of property right.
Comparative example
Comparison of therapeutic
One class new antitumoral reactive compound, they are contrasted with VX-680, and the compound ii b of invention is to human cervical carcinoma cell
HeLa, Non-small cell lung carcinoma cell A-549, tetra- kinds of tumour cells of human liver cancer cell HepG2 and human colon cancer cell LoVo
GIA is better than VX-680.Wherein compound ii d is to tetra- kinds of cancer cells of HeLa, A-549, HepG2 and LoVo
Inhibitory activity be respectively 38,32,44 and 6 times of VX-680.The suppression of chemical compounds I g and II d to Aurora A simultaneously
Activity is respectively 20 times and 32 times of VX 680.
Claims (6)
1. the BTAK inhibitor that a kind of stable nitrogen-oxygen free radical of structure with active anticancer such as formula I is marked,
Wherein：
R is ortho position chlorine, or meta chlorine, or contraposition chlorine, or vicinal hydroxyl groups, or meta-hydroxyl, or contraposition hydroxyl.
X is nitro, or fluorine, or chlorine, or bromine.
2. the BTAK inhibitor that a kind of stable nitrogen-oxygen free radical of structure with active anticancer such as formula II is marked,
Wherein：
R is ortho position chlorine, or meta chlorine, or contraposition chlorine.
X is nitro, or fluorine, or chlorine, or bromine.
3. the preparation method of compound described in claim 1, it is characterized in that with 2, the chloro- 5- nitro-pyrimidines of 4- bis-, or 2, the chloro- 5-FUs of 4- bis-, or 2, 4, 5- trichloropyrimidines, or 2, the chloro- 5- Bromopyrimidines of 4- bis- are raw material, first and o-chloraniline, or m-chloroaniline, or parachloroanilinum, or ortho-aminophenol, or a hydroxyanilines, or the reaction of para hydroxybenzene amine generates the chloro- 4- o-chloranilines -5- nitro-pyrimidines of 2-, or the chloro- 4- m-chloroanilines -5- nitro-pyrimidines of 2-, or the chloro- 4- parachloroanilinum -5- nitro-pyrimidines of 2-, or the chloro- 4- ortho-aminophenols -5- nitro-pyrimidines of 2-, or hydroxyanilines -5- nitro-pyrimidines between 2- chloro- 4-, or the chloro- 4- para hydroxybenzenes amine -5- nitro-pyrimidines of 2-, or the chloro- 4- o-chloranilines -5-FUs of 2-, or 2, the chloro- 4- o-chloranilines pyrimidines of 5- bis-, or the chloro- 4- o-chloranilines -5- Bromopyrimidines of 2-；Simultaneously the p-aminobenzoic acid with tertbutyloxycarbonyl protection is as raw material; with 4- amino -2 under conditions of 1- (3- dimethylamino-propyls) -3- ethyl-carbodiimide hydrochlorides and I-hydroxybenzotriazole are present; 2; 6; 6- tetramethyl piperidines nitrogen oxides is condensed, then deprotection base obtains final product p-benzoyl 4- amino -2 in the dichloromethane solution of trifluoroacetic acid, and 2; 6,6- tetramethyl piperidine nitrogen oxides；Last two kinds of compounds obtained above obtain the target compound representated by formula I under strong acid catalyst in the 2- positions generation substitution reaction of pyrimidine ring.
4. the preparation method of compound described in claim 2, it is characterized in that with 2, the chloro- 5- nitro-pyrimidines of 4- bis-, or 2, the chloro- 5-FUs of 4- bis- are raw material, first with p-benzoyl o-chloraniline, or p-benzoyl m-chloroaniline, or the reaction of p-benzoyl parachloroanilinum generates N- (the chloro- 5- nitro-pyrimidins -4 of 2-)-aminobenzoyl o-chloraniline, or N- (the chloro- 5- nitro-pyrimidins -4 of 2-)-aminobenzoyl m-chloroaniline, or N- (the chloro- 5- nitro-pyrimidins -4 of 2-)-aminobenzoyl parachloroanilinum, or N- (the fluoro- pyrimidines -4 of the chloro- 5- of 2-)-aminobenzoyl o-chloraniline；Simultaneously the p-aminobenzoic acid with tertbutyloxycarbonyl protection is as raw material; with 4- amino -2 under conditions of 1- (3- dimethylamino-propyls) -3- ethyl-carbodiimide hydrochlorides and I-hydroxybenzotriazole are present; 2; 6; 6- tetramethyl piperidines nitrogen oxides is condensed, then deprotection base obtains final product p-benzoyl 4- amino -2 in the dichloromethane solution of trifluoroacetic acid, and 2; 6,6- tetramethyl piperidine nitrogen oxides；Last two kinds of compounds obtained above obtain the target compound representated by formula II under strong acid catalyst in the 2- positions generation substitution reaction of pyrimidine ring.
5. application of the compound described in claim 1 or 2 as BTAK inhibitor.
6. the compound described in claim 1 or 2 applies the application in cancer therapy drug.
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN113773263A (en) * | 2020-06-10 | 2021-12-10 | 兰州大学 | 4-amine substituted phthalazinone aurora kinase B inhibitor and preparation and application thereof |
| WO2022270987A1 (en) * | 2021-06-24 | 2022-12-29 | (주) 업테라 | Aurka selective degradation inducing compound |
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|---|---|---|---|---|
| WO2008079719A1 (en) * | 2006-12-19 | 2008-07-03 | Genentech, Inc. | Pyrimidine kinase inhibitors |
| WO2012135641A2 (en) * | 2011-03-30 | 2012-10-04 | H. Lee Moffitt Cancer Center And Research Institute | Aurora kinase inhibitors and methods of making and using thereof |
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2015
- 2015-11-09 CN CN201510753988.3A patent/CN106674197A/en active Pending
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2008079719A1 (en) * | 2006-12-19 | 2008-07-03 | Genentech, Inc. | Pyrimidine kinase inhibitors |
| WO2012135641A2 (en) * | 2011-03-30 | 2012-10-04 | H. Lee Moffitt Cancer Center And Research Institute | Aurora kinase inhibitors and methods of making and using thereof |
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| FRANKLIN J. MOY ET AL.: "Novel Synthesis and Structural Characterization of a High-Affinity Paramagnetic Kinase Probe for the Identification of Non-ATP Site Binders by Nuclear Magnetic Resonance", 《JOURNAL OF MEDICINAL CHEMISTRY》 * |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN113773263A (en) * | 2020-06-10 | 2021-12-10 | 兰州大学 | 4-amine substituted phthalazinone aurora kinase B inhibitor and preparation and application thereof |
| CN113773263B (en) * | 2020-06-10 | 2023-09-19 | 兰州大学 | 4-amine substituted phthalazinone aurora kinase B inhibitor and preparation and application thereof |
| WO2022270987A1 (en) * | 2021-06-24 | 2022-12-29 | (주) 업테라 | Aurka selective degradation inducing compound |
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